Deciphering the family of ankyrin repeat-containing proteins of Plasmodium falciparum: a structural and functional perspective.

The invasion of host erythrocytes by Plasmodium falciparum is a fundamental step in its pathogenesis, relying on precisely regulated, transient interactions between parasite proteins and the host erythrocytes. Approximately 20% of eukaryotic proteins contain tandem repeat regions that majorly facilitate protein-protein interactions, with ankyrin (ANK) repeats being the most frequent, and are associated with various diseases. Despite significant advancements in understanding the importance of ANK repeats across various organisms, their roles within the phylum Apicomplexa, including P. falciparum, remain largely unexplored. This study focuses on the significance of ANK repeat proteins in P. falciparum growth and pathogenesis. Here, using in silico approaches, we deciphered the ANK repeat protein family in P. falciparum and elucidated the crystal structure of one of its members, PfANK_1. Co-immunoprecipitation assays revealed several potential interacting partners of PfANK_1, showing its involvement in the functioning of the PfClag9-PfRhopH complex. The stability and interactions of PfANK_1 within the complex were confirmed through multiple approaches, such as bio-layer interferometry (BLI), co-localization studies, and molecular dynamic simulations. Given the crucial role of the PfClag9-PfRhopH complex in erythrocyte invasion, PfANK_1 presents a promising druggable target. The structural analysis uncovered the binding interface and key interacting residues of PfANK_1 with the components of the PfClag9-PfRhopH complex. The work thus provides the first molecular and structural characterization of an ANK repeat protein in P. falciparum, offering new insights into the role of these proteins in the parasite life cycle and providing a potential platform for structure-function relationship-guided drug discovery.